Iron deficiency (ID) is the most prevalent single deficiency state on a worldwide basis. It is important economically because it diminishes the capability of individuals who are affected to perform physical labor, and it diminishes both growth and learning in children.
Absolute iron deficiency, with anemia or without anemia, and functional iron deficiency (FID) are high frequency clinical conditions, and these patients have iron deficient erythropoiesis. Absolute iron deficiency is defined as a decreased total iron body content. Iron deficiency anemia (IDA) occurs when iron deficiency is sufficiently severe to diminish erythropoiesis and cause the development of anemia. Functional iron deficiency describes a state where the total iron content of the body is normal or even elevated, but the iron is ‘locked away’ and unavailable for the production of red blood cells. This condition is observed mainly in patients with chronic renal failure who are on hemodialysis.
Iron status can be measured using hematological and biochemical indices. Each parameter of iron status reflects changes in different body iron compartments and is affected at different levels of iron depletion. Specific iron measurements include hemoglobin (Hgb), mean cell volume (MCV), hematocrit (Hct), erythrocyte protoporphyrin, plasma iron, transferrin, transferrin saturation levels (TSAT), serum ferritin (SF) and more recently soluble transferrin receptors (sTfR) and red-cell distribution width (RDW).
Typical values for normal iron status are SF 100±60 ng/ml and Hgb 12-17 g/dl for women and 14-19 g/dl for men. The typical values for latent iron deficiency are SF <22 ng/ml and Hgb normal to slightly low. The typical values for iron deficiency anemia are SF <22 ng/ml, Hgb for women <12 g/dl and for men <13 g/dl.
Hemoglobin (Hgb) has been used longer than any other iron status parameter. It provides a quantitative measure of the severity of iron deficiency once anemia has developed. Hemoglobin determination is a convenient and simple screening method and is especially useful when the prevalence of iron deficiency is high, as in pregnancy or infancy. The limitations of using hemoglobin as a measure of iron status are its lack of specificity (as factors such as B12 or folate deficiency, genetic disorders and chronic infections can limit erythropoiesis) and its relative insensitivity due to the marked overlap in values between normal and iron deficient populations. To identify iron deficiency anemia, hemoglobin is measured together with more selective measurements of iron status.
A reduction in mean cell volume (MCV) occurs when iron deficiency becomes severe, at about the same time as anemia starts to develop. It is a fairly specific indicator of iron deficiency once thalassemia and the anemia of chronic disease have been excluded. A cut-off value of 80 fl is accepted as the lower limit of normal in adults.
The red-cell distribution width (RDW) has been used recently in combination with other parameters for the classification of anemias. It reflects the variation in the size of the red cells and can be used to detect subtle degrees of anisocytosis. RDW is computed directly form the RBC histogram. Two different calculated values have been provided on hematology analyzers. The RDW-CV is measured as a ratio of the width of the distribution curve at one standard deviation divided by the MCV. The RDW-SD is a direct measurement of the distribution width at the 20% frequency level. Normally, the size distribution curve for red blood cells is quite symmetrical, with an RDW-CV value of 10±1.5% and an RDW-SD of 42±5 (fl). A high RDW, which means a greater variation in cell size, is caused by either the appearance of macrocytic or microcytic cells. An elevated red-cell distribution width appears to be the earliest hematological manifestation of iron deficiency.
The most commonly used iron status parameters at present are transferrin saturation (TSAT) and serum ferritin (SF). However, both are indirect measures of iron status. Transferrin is a transport protein that contains two iron binding sites by which it transports iron from storage sites to erythroid precursors. TSAT (i.e., the percentage of total binding sites that are occupied by iron) is a measure of iron that is available for erythropoiesis. TSAT is calculated by dividing the serum iron by the total iron binding capacity (TIBC), a measurement of circulating transferrin, and multiplying by 100. Ferritin is a storage protein that is contained primarily within the reticuloendothelial system (RES), with some amounts released in the serum. Under conditions of iron excess, ferritin production increases to offset the increase in plasma iron. The level of ferritin in the serum, therefore, reflects the amount of iron in storage.
For patients with chronic kidney disease, absolute iron deficiency may be diagnosed when TSAT is <20% and SF is <100 ng/ml. Functional iron deficiency may be more difficult to diagnose since iron status parameters may indicate adequate iron stores. There are different criteria in defining FID, one of them is published by the Kidney Disease Outcomes Quality Initiative- K/DOQI (Eknoyan G, et al. Continuous quality improvement: DOQI becomes K/DOQI and is updated. National Kidney Foundation's Dialysis Outcomes Quality Initiative. Am J Kidney Dis., 2001 January; 37(1):179-194), as shown in the following table.
Definition of Functional Iron Deficiency (FID) and Absolute IronDeficiency (AID) by Kidney Disease Outcomes, QualityInitiative K/DOQI (U.S.A)Ferritin μg/L<100100-800TSAT <20%AIDTSAT 20%-50%FID
The limitations of using transferrin saturation reflect those of serum iron, i.e., wide diurnal variation and low specificity. TSAT is also reduced in inflammatory disease. Transferrin saturation is commonly used in population studies combined with other indicators of iron status. On the other hand, as ferritin is an acute phase reactant, its serum levels may be elevated in the presence of chronic inflammation, infection, malignancy and liver disease. Alcohol consumption has also been suggested to independently raise serum ferritin.
Recently, several new red blood cell and reticulocyte parameters have been reported having utilities in detection of iron deficiency and functional iron deficiency. Two of the parameters are hypochromic red cells % (referred to as % Hypo) and CHr (reticulocyte hemoglobin content) reported by the Bayer ADVIA® 120 hematology analyzer (Thomas C. et al. Biochemical Markers and Hematologic Indices in the Diagnosis of Functional Iron Deficiency. Clinical Chemistry 48:7, 1066-1076, 2002). CHr is defined by the formula (CHr=MCVr×CHCMr), wherein MCVr is the mean reticulocyte cell volume and CHCMr is the mean hemoglobin concentration of reticulocytes which is obtained by an optical cell-by-cell hemoglobin measurement.
Reticulocytes are immature red blood cells with a life span of only 1 to 2 days. When these are first released from the bone marrow, measurement of their hemoglobin content can provide the amount of iron immediately available for erythropoiesis. A less than normal hemoglobin content in these reticulocytes is an indication of inadequate iron supply relative to demand. The amount of hemoglobin in these reticulocytes also corresponds to the amount of hemoglobin in mature red blood cells. CHr has been evaluated recently in numerous studies as a test for iron deficiency and functional iron deficiency and has been found to be highly sensitive and specific. However, exact threshold values have not been established, as the threshold values vary depending on the laboratory and instrument used.
Epoetin is effective in stimulating production of red blood cells, but without an adequate iron supply to bind to heme, the red blood cells will be hypochromic, i.e., low in hemoglobin content. Thus, in states of iron deficiency, a significant percentage of red blood cells leaving the bone marrow will have a low hemoglobin content. By measuring the percentage of red blood cells with hemoglobin content <28 g/dl, iron deficiency can be detected. Hypochromic red cells percentages >10% have been correlated with iron deficiency. % Hypo is reported by Bayer ADVIA 120 hematology analyzer based on the optical cell-by-cell hemoglobin measurement.
Two other parameters have been reported recently correlating to %Hypo and CHr are RBC-Y and RET-Y reported by the SYSMEX® XE-2100 hematology analyzer (Machin S.J. et al. Functional Iron Deficiency and New Red Cell Parameters on the Sysmex XE-2100, ISLH 2001 Industry-Sponsored Workshops, ISLH XIVth International Symposium. 2001). RBC-Y is the mean value of the forward light scatter histogram within the mature erythrocyte population, and RET-Y is the mean value of the forward light scatter histogram within the reticulocyte population obtained in a reticulocyte measurement on the SYSMEX® XE-2100 hematology analyzer.
Another parameter that has been used previously for detection of iron deficiency is the ratio of MRV/MCV, or In (MRV/MCV) (referred to as dR), wherein MRV is the mean reticulocyte cell volume and MCV is the mean red blood cell volume. Typically, the ratio of MRV/MCV >1.35 is considered as the indication of iron deficiency.
Furthermore, in a different aspect from assisting diagnosis and treatment of diseases, it is desirable to detect certain hematological conditions before the occurrence of the diseases. It is well known that latent iron deficiency (LID) has a high frequency in fertile women, due to menstruation and sometimes due to poor diet. Latent iron deficiency refers to the presence of iron deficiency but not yet anemia. Also well known is the relatively high frequency of latent functional iron deficiency (LFID) and hemochromatosis (HEM) in the general population. Latent functional iron deficiency refers to the pre-anemic stage of functional Iron deficiency. On the other hand, hemochromatosis, the most common form of iron overload disease, is an inherited disorder that causes the body to absorb and store too much iron. The extra iron builds up in organs and causes damages to the organs. Without treatment, the disease can cause these organs to fail. Clinically, it is important to detect these conditions early in order to provide preventative treatments.
It is desirable to be able to detect iron deficiency, such as absolute iron deficiency, latent iron deficiency, functional iron deficiency and latent functional iron deficiency, using existing hematology parameters reported on an automated hematology analyzer during a blood analysis routinely performed on the instrument, which can assist in early detection of the clinical conditions, without additional cost.